Purified vaccina virions contain a large number of enzymatic activities. Many of those activities which have a defined function deal with the biogenesis of mature viral mRNA. These functions can be studied in a relatively clean system in vitro. This, coupled with the essentially cytoplasmic nature of the viral infectious cycle in vivo, make vaccinia virus an ideal system for studying the complexities involved in the expression and regulation of viral genetic information. We would like to pursue our observation of the synthesis of small RNA molecules by vaccinia virus. We consider it of great interest that these small RNA's (ranging in size from 20 to several hundred bases) are capped and methylated at the 5'-terminus and polyadenylated at the 3' terminus. As reagents for measuring promoters they will be used to study the expression of viral genes in vitro and their temporal regulation and expression in vivo. The possibility that these small RNA molecules may be involved along with the high molecular weight RNA synthesized by the virus in the maturation of viral monocistronic-sized messages will be investigated. Studies on the endoribunuclease associated with vaccinia virions has shown a great deal of specificity with vaccinia RNA's. The solubilized enzyme(s) has been used to generate limited cleavage products from vaccinia RNA as well as genomic RNA from influenza and polioviruses. With this regard, our goal is to purify the endoribonuclease(s) to study the physico-chemical nature of the enzyme(s) and to analyze the basis for the apparent limited cleavage products from these RNA substrates. If base sequence recognition were required for this endoribonuclease(s), it would be quite exciting. Analysis of the DNA genome of the virus is being pursued with the intention of understanding gene expression and regulation as well as utilizing the DNA fragments as a tool for studying the expression and biogenesis of viral mRNA.